The leading cause of dementia and the fourth leading cause of death in the developed world is Alzheimer disease, which afflicts an estimated 10% of the population over 65 years of age in the United States. The disease manifests itself as insidious memory loss, cognitive decline, and personality changes that result in loss of functional ability over the course of a decade. In their debilitated state, patients usually retain only vegetative neurologic function, and succumb to secondary infections.
Alzheimer disease is characterized by certain neuropathological lesions, including intracellular neurofibrillary tangles and extracellular parenchymal and cerebrovascular amyloid. The principal component of the amyloid deposits is a protein designated .beta./A4 amyloid (4, 5), a .about.4 kDa polypeptide arising from cleavage of the amyloid precursor protein (APP), see Goldgaber et al., Science, 235, pp. 887-880 (1987). APP exists as three major transmembrane isoforms (APP.sub.695, APP.sub.751, and APP.sub.770) that result from alternative splicing of a single primary transcript (FIG. 1A), see Kang et al., Nature (London), 325, pp. 733-736 (1987). Proteolytic processing of APP leads to cleavage within the .beta./A4 domain and precludes amyloidogenesis. The biochemical defect responsible for amyloid production in Alzheimer disease might therefore involve either a deficiency in normal proteolysis or excessive activity of an alternative pathway. It is noteworthy that two types of inherited cerebral amyloidoses--hereditary cerebral hemorrhage with amyloidosis (Dutch type) and familial early onset Alzheimer disease--are associated with mutations in the coding sequence of APP near the .beta./A 4-amyloid domain. Alzheimer disease is characterized by abnormal protein phosphorylation and altered protein catabolism. From the work of several laboratories, altered protein phosphorylation has been implicated in the formation of the intracellular neurofibrillary tangles found in Alzheimer disease. However, a role for protein phosphorylation in the catabolism of the .beta./A4 protein precursor (.beta.APP) has not been demonstrated.
A central feature of the pathology of Alzheimer disease is the deposition of amyloid protein within plaques. The 4 kDa amyloid protein (also referred to as A4 (APC, .beta.-amyloid or BAP) is a truncated form of the larger amyloid precursor protein (APP) which is encoded by a gene localized on chromosome 21 (Goldgaber et al., 1987, Science, 235:877-880; Kang et al., 1987, Nature. 325:733-736) Jenkins et al., 1988, Biochem. Biophys. Res. Commun., 151:1-8; Tanzi et al., 1987, Science. 235:880-885). Genetic linkage analysis, using DNA probes that detect restriction fragment length polymorphisms (RFLPS, Botstein et al., 1980, Am. J. Hum. Genet., 32:314-331), has resulted in the localization of a candidate gene (FAD, familia Alzheimer disease) on human chromosome 21 in families with high frequencies of Alzheimer's disease (St. George-Hyslop et al., 1987, Science, 235:885-890). However, the FAD locus has not been localized precisely, and very little is known about its function. Initial studies of individuals with Down's Syndrome (DS), caused by trisomy of chromosome 21, indicate that these individuals develop Alzheimer-like pathology beyond the second decade of life. However, analysis of multiple Alzheimer pedigrees revealed that the APP gene does not segregate with familial Alzheimer disease (Van Broackhoven et al., 1987, Nature. 328:153-155; Tanzi et al., 1987, Nature, 329:156-157). Furthermore, two recent studies with new families demonstrated the absence of a linkage of chromosome 21 markers to familial Alzheimer disease (Schellenberg et al , 1988, 241:1507-1510; Rosea et al., 1988, Neurology, 38:173).
Age, genetic elements, and, possibly environmental factors appear to contribute to cellular pathology of Alzheimer's disease. A fundamental but unanswered question in the pathogenesis of Alzheimer disease is the relationship between abnormalities of neurons and the deposition of amyloid. Specifically, the cellular origin of pathological events leading to the deposition of amyloid fibrils adjacent to some areas of the blood-brain barrier (cerebrovascular amyloid) and in the proximity of nerve terminals (neuritic plaques) in specific brain regions as well as extracellular amyloid in plaques cores is not known. Glenner and Wong have described the purification and characterization of meningeal amyloid from both brains of individuals with Alzheimer disease (Glenner and Wong, 1984, Biochem. Biophys. Res. Commun., 120:885-890) or Down's Syndrome (Glenner & Wong, 1984, Biochem. Biophys. Res. Comm., 122:1131-1135) and determined the N-terminal peptide sequences. Among 24 residues analyzed, the two amyloid peptides showed only one difference, namely at amino acid position 11 (glutamine in Alzheimer disease amyloid, versus glutamic acid in Down's Syndrome amyloid) among 24 residues analyzed. Subsequent studios of amyloid from Alzheimer brain plaque cores revealed amino acid sequences identical to the reported Down's Syndrome cerebrovascular amyloid data (Masters et al, 1985, Proc. Natl. Acad. Sci., U.S.A., 82:4245-4249). Copy-DNA analysis of APP transcripts form both normal tissue and Alzheimer brain material demonstrated the presence of the codon for glutamic acid at this position (Kang et al., 1987, supra; Goldgaber et al, 1987, supra; Robakis et al., supra; Tanzi et al., 1987, Science, 235:880-884; Zain et al., 1988, Proc. Natl. Acad. Sci., U.S.A., 85:929-933; Vitek et al., 1988, Mol. Brain Res., 4:121-131).
Cerebral .beta./A4-amyloidosis is characteristic of several apparently related human conditions: Alzheimer's disease, Down's syndrome, hereditary cerebral hemorrhage with amyloidosis, Dutch type, and, to a lesser extent, normal aging. Like other organ-specific amyloidoses, the pathology of these conditions includes the accumulation of extracellular protein precipitates which form .beta.-pleated sheets and bind Congo red dye (Cotran et al., "Robbin's Pathologic Basis of Disease," 4th ed., WB Saunders, Pa., 1984). Unlike other amyloidoses, however, in which the deposited peptide seems clearly derived from a hydrophilic domain in a circulating serum precursor, deposited .beta./A4-amyloid includes a portion of the intramembranous domain of an integral transmembrane protein, the Alzheimer .beta./A4 amyloid precursor protein (APP). This raises several novel questions about how APP is processed within the cell and about how .beta./A4 is released into the extracellular space. Synthetic .beta./A4 spontaneously aggregates and precipitates (15), thus demonstrating that the primary amino acid sequence of this domain of APP encodes sufficient structural information to determine its amyloidogenicity.